Development of FRET-based cap-snatching endonuclease assay

Viruses belonging to the order <i>Bunyavirales</i>, including Hantaviruses, Crimean-Congo hemorrhagic fever virus, Rift Valley fever virus, Severe Fever with Thrombocytopenia Syndrome Virus, and La Crosse encephalitis virus, cause severe human illnesses with mortality rates in certain outbreaks reaching 50%, 10%-40%, 10%-20%, 6%-30%, and 1%, respectively. Currently, there are no Food and Drug Administration-approved vaccines or antiviral therapeutics available for these viruses. The highly efficient and cost-effective fluorescence resonance energy transfer-based <i>in vitro</i> endonuclease assay, having a quantitative fluorescence readout, can be optimized for high-throughput screening of chemical libraries to identify chemical inhibitors for the Bunyavirus cap-snatching endonuclease. The assay will be of critical importance for antiviral drug discovery against numerous negative-strand RNA viruses using cap-snatching mechanism for transcription initiation and replication of the RNA genome.